Functional genomics requires reliable methods to investigate the consequences of disrupting the gene function. Since its discovery in C .elegans, RNA interference (RNAi) has become an invaluable resource to tackle gene function in cell culture, mammals and invertebrate model systems. However, but it has proven a major challenge to apply this technique in zebrafish and Xenopus. In this proposal we will apply our current knowledge in the microRNA (miRNA) field to develop several strategies that improve RNAi. This proposal will provide specific rules to the scientific community to improve RNAi in zebrafish to allow the functional characterization of the zebrafish genes. In the future, the resuls derived from this project will establish a frame work to understand the structure and the function of vertebrate genes. PUBLIC HEALTH RELEVANCE: Understanding the function of genes requires reliable methods to block gene expression. Over the last decade, it has been discovered that double strand RNA can be used to shut down expression of genes (RNA interference) however this technique is not available in zebrafish. This proposal aims to understand the steps required to apply this powerful technique to zebrafish, what will help us understand the function of vertebrate genes in development and disease.